CN110833568B - Use of durian heart skin extract for improving muscle fatigue - Google Patents
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Abstract
The invention relates to application of durian heart-skin extract, in particular to application of durian heart-skin extract in improving muscle fatigue. The durian heart skin extract provided by the invention can be used for promoting the proliferation of skeletal muscle cells and improving the activity of the skeletal muscle cells so as to improve muscle fatigue. The durian heart skin extract prepared by the invention can also promote the expression of SOD3 or NADSYN1 genes and inhibit the expression of PARP1 or PARP2 genes, thereby increasing the generation of NAD +, improving the activity of mitochondria, increasing physical strength and energy and improving the problem of muscle fatigue.
Description
Technical Field
The invention relates to an application of a durian heart skin extract, in particular to an application of a durian heart skin extract in improving muscle fatigue and regulating and controlling related gene expression.
Background
In modern multi-element society, everyone strives at work stations for swabbing, and in order to achieve good work performance, the person needs to have good professional ability and healthy body. Therefore, how to maintain the health of the body has become an important target pursued by modern people. In order to maintain health, the most important people have too much choice of diet besides exercise, so that the most beneficial people are well-benefited if foods with various nutrient components and capable of improving physical strength can be found.
In the general nutritional product market, there are many foods or drinks with refreshing components, but there may be many unknown components contained therein, and there may be effects of temporarily increasing physical strength and refreshing, but there may be many substances harmful to the body, so it is an important choice to select a food or a composition having an effective component for increasing physical strength without producing other harmful effects.
Durian (Durio zibethinus) is known as "fruit king" in Thailand, Malaysia, India and the like, and its unique odor is often unacceptable for many consumers. The fruits of durian are mostly oblong or round, the color is brown with green color, the pulp is light yellow, and the shell is all over with convex thorns. The durian pulp contains a lot of nutrient components, besides various vitamins, fat, calcium, iron, phosphorus, saccharide and other components, also contains various amino acids and dietary fibers, so the durian pulp has the efficacies of enhancing immunity, promoting gastrointestinal motility and the like. However, there is a heart-skin portion formed by separating the flesh of durian, namely, durian white flesh, besides the flesh and hard shell, and there is no research on any component or efficacy at present, and it is generally discarded with the shell after the flesh is taken out, so that if it can be recycled or further ingredients beneficial to human health are found, it can also bring benefits to both environment protection and human health.
Disclosure of Invention
One of the objectives of the present invention is to provide an application of durian heart-skin extract for improving muscular fatigue, which promotes the proliferation of skeletal muscle cells, increases the activity of skeletal muscle cells, and enhances physical strength and energy.
In an embodiment of the present invention, the durian heart-skin extract can be obtained by water extraction. In one embodiment, the durian heart-skin extract can be obtained by extraction at 50-100 ℃.
In one embodiment of the present invention, the dosage of the durian heart-bark extract can be about 0.2-0.3 mg/ml, preferably about 0.25 mg/ml.
In one embodiment of the present invention, the durian heart-skin extract can be used to promote the expression of gene SOD3 or NADSYN 1.
In another embodiment of the present invention, the durian heart bark extract can be used to inhibit the expression of the genes PARP1 or PARP 2.
In yet another embodiment of the present invention, the durian heart skin extract can be further added to food, health food or dietary supplement. In addition, the durian heart-skin extract can be further used for preparing a pharmaceutical composition.
The following examples are presented to illustrate the present invention and are not to be construed as limiting the scope of the invention, which is intended to be limited only by the appended claims.
Drawings
FIG. 1 is a graph comparing the results of durian heart skin extract on the proliferative effects of skeletal muscle cells according to an embodiment of the present invention;
FIG. 2 is a graph comparing the results of durian heart skin extract effects on skeletal muscle cell activity according to embodiments of the present invention;
FIG. 3 is a graph showing the results of the effect of durian heart-skin extract on PARP1 and PARP2 gene expression;
FIG. 4 is a graph showing the effect of the durian heart-skin extract on the expression of SOD3 and NADSYN1 genes.
Detailed Description
Example 1 preparation of durian heartbark extract
Taking out a proper amount of durian heart skin, and cleaning for later use. Mixing durian heart skin with water at a liquid-solid ratio of 2-10: 1-5, homogenizing, and performing extraction reaction at 50-100 deg.C for 0.5-3 hr. After the reaction, the mixture was cooled to room temperature and filtered through a 400-mesh screen. And then carrying out reduced pressure concentration on the filtrate at the temperature of 45-70 ℃, and concentrating to obtain the durian heart skin extract (extract liquor).
Example 2 skeletal muscle cell proliferation assay
Mouse skeletal muscle cells (C2C12, ATCC CRL-1772) were prepared and cultured in a cell culture medium [ Dulbecco's Modified Eagle's Medium (DMEM), 1% Penicillin-streptomycin (Gibco), 10% fetal bovine serum (Gibco) ]. A96-well plate was prepared, 5000 cells were implanted in each well, and then cultured at 37 ℃ for 2 hours.
The test was performed in four groups, where group A was blank, group B was control, 10% fetal bovine serum was added, and C, D were test groups to which the durian heart-skin extract prepared in example 1 was added at concentrations of 0.25mg/ml and 0.125mg/ml, respectively. The assay was performed with a cell proliferation ELISA kit (Roche, 11647229001). First of all. Each group was added 100. mu.l of the test component to each well, and 100. mu.l of BrdU labeling solution (containing 5-bromo-2' -deoxyuridine) at a concentration of 100. mu.M was added thereto, and reacted at room temperature for 24 hours.
The supernatant was then removed and 200. mu.l of FixDenat solution was added to each well and allowed to act at room temperature for 30 minutes. The FixDenat solution was then removed and washed once with 1 XPBS (Gibco), and 100. mu.l of anti-BrdU-POD solution was added to each well and allowed to stand at room temperature for 90 minutes. After removing the antibody conjugate, each well was washed three times with 200-300 μ l of washing solution. Then, after removing the washing solution, 100. mu.l of the substrate solution was added to each well and allowed to act at room temperature for 5 to 30 minutes. Finally, 25. mu.l of 1M sulfuric acid was added to each well and allowed to act for about 1 minute with shaking at 300 rpm. The reacted solution was then measured for absorbance at a wavelength of 450 nm. The measured values were analyzed for statistical significance between the two values using the Student's t-test using microsoft EXCEL software, and the results are shown in fig. 1. Wherein, in contrast to the blank group, denotes a P value < 0.05, a P value < 0.01, and a P value < 0.001.
As can be seen from the results shown in fig. 1, under the effect of the durian heart bark extract containing the durian heart bark extract of the present invention at a concentration of 0.25mg/ml, the detected BrdU amount was increased by 32% compared to the control group, i.e., the number of skeletal muscle cells was increased by 32%, thus demonstrating that the durian heart bark extract of the present invention does have the effect of promoting the proliferation of skeletal muscle cells.
Example 3 skeletal muscle cell mitochondrial Activity assay
Mouse skeletal muscle cells (C2C12, ATCC CRL-1772) were prepared and cultured in a cell culture medium [ Dulbecco's Modified Eagle's Medium (DMEM), 1% Penicillin-streptomycin (Gibco), 10% fetal bovine serum (Gibco) ]. A96-well plate was prepared, 5000 cells were implanted in each well, and then cultured at 37 ℃ for 2 hours.
The test was performed in three groups, where group a was a blank and group B, C was a test group to which 0.25mg/ml of the durian heart-skin extract prepared in example 1 was added, for 2 and 24 hours respectively. The method for performing the activity test of the mitochondria in the test is to use the trypsin as the cells attached to the culture dishThe enzyme (trypsin) was excised, neutralized with medium and centrifuged to remove the supernatant as BDTMThe MitoScreen (JC-1) mitochondrial specific reagent was stained for 15min, washed twice with assay buffer (assay buffer) attached to the reagent, and subjected to mitochondrial activity analysis using a BD Accuri C6 Plus flow cytometer.
As can be seen from the results in fig. 2, the mitochondrial activity of the durian heart bark extract containing the durian heart bark extract of the present invention at a concentration of 0.25mg/ml was greatly increased by about 51 to 66% compared to the blank group in both 2 hours and 24 hours, which shows that the durian heart bark extract of the present invention can increase the cell activity of skeletal muscle cells and has the effect of improving muscle endurance and energy.
Example 4 testing of mitochondrial Activity-associated Gene expression
Mouse skeletal muscle cells (C2C12, ATCC CRL-1772) were prepared and cultured in a cell culture medium [ Dulbecco's Modified Eagle's Medium (DMEM), 1% Penicillin-streptomycin (Gibco), 10% fetal bovine serum (Gibco) ]. The test was divided into control and test groups, in which the durian carpel extract prepared in example 1 was added at an action concentration of 0.125mg/ml and cultured at 37 ℃ for 6 and 24 hours, respectively, and then examined to analyze the expression status of genes related to mitochondrial activity, particularly genes related to regulation of NAD +.
The cells after the above-mentioned reaction were collected, and their RNAs were extracted using an RNA extraction kit (Geneaid), and these RNAs were Reverse-transcribed into cDNAs using Reverse transcriptase (SuperScript III Reverse transcriptase Transcriptase, Invitrogen). Then, qPCR (KAPA CYBR FAST qPCR Kits, KAPA Biosystems) was performed using the primers listed in table 1 using a Real-Time polymerase chain reaction system (ABI Step One Plus Real-Time PCR system) to quantify the expression of the following genes: PARP1, PARP2, SOD3 and NADSYN1 (expression of ACTB gene is used as internal control group). The relative quantitative analysis of gene expression is 2-ΔΔCtMethods, and statistical significance between values was analyzed using Excel software with student assay. The results of gene expression are shown in FIGS. 3 and 4. Wherein, represents a P value < 0.05, represents a P value < 0.01, and represents a P value < 0.001.
TABLE 1
From the results of fig. 3, it is known that the expression level of PARP1 gene is reduced by about 30-50% and the expression level of PARP2 gene is reduced by about 40% under the action of the extract containing the durian heart skin according to the embodiment of the present invention. Since the PARP1 and PARP2 genes inhibit the production of NAD +, the expression of the genes is reduced, the production of NAD + is increased, and the mitochondria maintain better activity, thereby having the effect of increasing physical strength.
On the other hand, as shown in fig. 4, the expression of SOD3 gene was greatly increased by about 60% to 270% under the action of the durian heart-skin extract containing the example of the present invention, and especially after 24 hours of action of the durian heart-skin extract, the expression of SOD3 gene was almost 4 times as high as the original expression. In addition, under the action of the durian heart-skin extract, the expression of the NADSYN1 gene can be increased by about 40%, the generation of NAD + is increased, related downstream genes are activated, the mitochondrial activity can be improved, and the effects of resisting aging and improving energy are achieved.
The above experiments show that the durian heart skin extract of the embodiment of the invention can promote the proliferation of skeletal muscle cells, improve the activity of the cells, and simultaneously regulate the expression of NAD + genes, so that the generation of NAD + is increased, thereby having the effects of improving muscle endurance, physical strength and energy.
In addition, the durian heart skin extract provided by the embodiment of the invention can be further added into food, health-care food or dietary supplement, or further applied to products such as cosmetics or maintenance products. Meanwhile, the durian heart-skin extract of the embodiment of the invention can be prepared into a pharmaceutical composition, and the pharmaceutical composition can be further added with carriers or other auxiliary agents well known in the technical field. The dosage form of the pharmaceutical composition may be, but is not limited to, a solution, a capsule, or a tablet.
Claims (11)
1. Non-therapeutic use of a durian heart skin extract for improving muscle fatigue by promoting skeletal muscle cell proliferation or increasing skeletal muscle cell mitochondrial activity.
2. Use according to claim 1, wherein said durian heart-skin extract is obtained by aqueous extraction.
3. Use according to claim 2, wherein the durian heart-skin extract is obtained by extraction at 50-100 ℃.
4. Use according to claim 1, wherein the dosage of durian heart-bark extract is 0.2-0.3 mg/ml.
5. Use according to claim 4, wherein the dosage of said durian heart-bark extract is 0.25 mg/ml.
6. Use according to claim 1, wherein the durian heart-bark extract is to promote genesSOD3 Or NADSYN1The expression of (1).
7. Use according to claim 1, wherein the durian heart-bark extract is used to inhibit genesPARP1OrPARP2The expression of (1).
8. Use according to claim 6 or 7, wherein the dosage of said durian heart bark extract is 0.1-0.3 mg/ml.
9. Use according to claim 1, wherein the durian heart-skin extract is further added to a food product.
10. Use according to claim 1, wherein the durian heart-skin extract is further added to health food.
11. Use according to claim 1, wherein the durian heart-skin extract is further added to a dietary supplement.
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| CN101244095A (en) * | 2007-06-07 | 2008-08-20 | 东莞市竟恒流通研究所 | New purpose of durio zibethinus L. shell extract |
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| CN107128650A (en) * | 2017-06-27 | 2017-09-05 | 盐城永悦新材料有限公司 | A kind of thermosetting powder coating double-screw additive feeding machine |
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| CN101244095A (en) * | 2007-06-07 | 2008-08-20 | 东莞市竟恒流通研究所 | New purpose of durio zibethinus L. shell extract |
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| Title |
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| 榴莲皮提取物抗炎作用研究;谢果等;《广州中医药大学学报》;20150120;第32卷(第1期);文章第130页摘要，左栏第1段，第131页左栏1.4 * |
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